The invention relates to a method for determining the concentration of fibrinogen in blood plasma using thrombin as a reagent.
Prior methods of using thrombin to measure fibrinogen concentration, include the Clauss method which is based on measuring the time it takes for a plasma-thrombin reaction to occur (clotting time) and the ACL3 fibrinogen assay. The Clauss method is described in Manual of Hemostasis and Thrombosis, ed. 3, by Arthur R. Thompson and Laurence A. Harker, Appendix A, p. 179 (1983) and in Gerrinnungs physiologische schnell Methode zur Bestimmung des Fibrinogens by A. Clauss, Acta Haematol, 17:237 (1957). The ACL3 method is described in Method for the Determination of Functional (Clottable) Fibrinogen by the New Family of ACL Coagulometers by E. Rossi, P. Mondonico, A. Lomabardi, L. Preda, Thrombosis Research 52; 453-469 (1988). These methods rely on the measurement of a relevant parameter such as clotting time or changes in optical transmission and on multiple dilutions of a calibrator plasma to compensate conditions of the instrument and reagent at a given time. Using calibrator plasmas (i.e. plasma having known fibrinogen concentrations) "standard lines" or "calibration curves" must be constructed repeatedly whenever conditions warrant. In the determination of fibrinogen concentration of an unknown sample, the relevant quantity, such as clotting time, is measured and the concentration is then "read" from the standard curve. This process can involve considerable calculation, and is often tedious and time consuming.
Another deficiency of these prior methods is that the relevant quantity being measured is often instrument dependent, as well as reaction dependent. For example, if the instrument used to measure the relevant parameter employs an electro-optical system in which scattered or transmitted light is detected, the value obtained from the measurement will depend on the signal level measured by the optical sensor, which in turn depends on the amount of light incident on the reaction vessel as well as the electronic gains used in association with the optical sensor. The values of these quantities do not remain constant in time, nor do they remain constant from channel to channel or instrument to instrument.